The 10th Biennial Hatter Cardiovascular Institute workshop: cellular protection—evaluating new directions in the setting of myocardial infarction, ischaemic stroke, and cardio-oncology

Due to its poor capacity for regeneration, the heart is particularly sensitive to the loss of contractile cardiomyocytes. The onslaught of damage caused by ischaemia and reperfusion, occurring during an acute myocardial infarction and the subsequent reperfusion therapy, can wipe out upwards of a billion cardiomyocytes. A similar program of cell death can cause the irreversible loss of neurons in ischaemic stroke. Similar pathways of lethal cell injury can contribute to other pathologies such as left ventricular dysfunction and heart failure caused by cancer therapy. Consequently, strategies designed to protect the heart from lethal cell injury have the potential to be applicable across all three pathologies. The investigators meeting at the 10th Hatter Cardiovascular Institute workshop examined the parallels between ST-segment elevation myocardial infarction (STEMI), ischaemic stroke, and other pathologies that cause the loss of cardiomyocytes including cancer therapeutic cardiotoxicity. They examined the prospects for protection by remote ischaemic conditioning (RIC) in each scenario, and evaluated impasses and novel opportunities for cellular protection, with the future landscape for RIC in the clinical setting to be determined by the outcome of the large ERIC-PPCI/CONDI2 study. It was agreed that the way forward must include measures to improve experimental methodologies, such that they better reflect the clinical scenario and to judiciously select combinations of therapies targeting specific pathways of cellular death and injury

Generation of a Homozygous Transgenic Rat Strain Stably Expressing a Calcium Sensor Protein for Direct Examination of Calcium Signaling

In drug discovery, prediction of selectivity and toxicity require the evaluation of cellular calcium homeostasis. The rat is a preferred laboratory animal for pharmacology and toxicology studies, while currently no calcium indicator protein expressing rat model is available. We established a transgenic rat strain stably expressing the GCaMP2 fluorescent calcium sensor by a transposon-based methodology. Zygotes were co-injected with mRNA of transposase and a CAG- GCaMP2 expressing construct, and animals with one transgene copy were pre-selected by measuring fluorescence in blood cells. A homozygous rat strain was generated with high sensor protein expression in the heart, kidney, liver, and blood cells. No pathological alterations were found in these animals, and fluorescence measurements in cardiac tissue slices and primary cultures demonstrated the applicability of this system for studying calcium signaling. We show here that the GCaMP2 expressing rat cardiomyocytes allow the prediction of cardiotoxic drug side-effects, and provide evidence for the role of Na+/Ca2+ exchanger and its beneficial pharmacological modulation in cardiac reperfusion. Our data indicate that drug-induced alterations and pathological processes can be followed by using this rat model, suggesting that transgenic rats expressing a calcium-sensitive protein provide a valuable system for pharmacological and toxicological studies

Effect of COMBinAtion therapy with remote ischemic conditioning and exenatide on the Myocardial Infarct size: a two-by-two factorial randomized trial (COMBAT-MI)

Remote ischemic conditioning (RIC) and the GLP-1 analog exenatide activate different cardioprotective pathways and may have additive effects on infarct size (IS). Here, we aimed to assess the efficacy of RIC as compared with sham procedure, and of exenatide, as compared with placebo, and the interaction between both, to reduce IS in humans. We designed a two-by-two factorial, randomized controlled, blinded, multicenter, clinical trial. Patients with ST-segment elevation myocardial infarction receiving primary percutaneous coronary intervention (PPCI) within 6 h of symptoms were randomized to RIC or sham procedure and exenatide or matching placebo. The primary outcome was IS measured by late gadolinium enhancement in cardiac magnetic resonance performed 3–7 days after PPCI. The secondary outcomes were myocardial salvage index, transmurality index, left ventricular ejection fraction and relative microvascular obstruction volume. A total of 378 patients were randomly allocated, and after applying exclusion criteria, 222 patients were available for analysis. There were no significant interactions between the two randomization factors on the primary or secondary outcomes. IS was similar between groups for the RIC (24 ± 11.8% in the RIC group vs 23.7 ± 10.9% in the sham group, P = 0.827) and the exenatide hypotheses (25.1 ± 11.5% in the exenatide group vs 22.5 ± 10.9% in the placebo group, P = 0.092). There were no effects with either RIC or exenatide on the secondary outcomes. Unexpected adverse events or side effects of RIC and exenatide were not observed. In conclusion, neither RIC nor exenatide, or its combination, were able to reduce IS in STEMI patients when administered as an adjunct to PPCI

Ischaemic conditioning and targeting reperfusion injury: a 30 year voyage of discovery

To commemorate the auspicious occasion of the 30th anniversary of IPC, leading pioneers in the field of cardioprotection gathered in Barcelona in May 2016 to review and discuss the history of IPC, its evolution to IPost and RIC, myocardial reperfusion injury as a therapeutic target, and future targets and strategies for cardioprotection. This article provides an overview of the major topics discussed at this special meeting and underscores the huge importance and impact, the discovery of IPC has made in the field of cardiovascular research

Lack of SMARCB1 expression characterizes a subset of human and murine peripheral T-cell lymphomas.

Peripheral T-cell lymphoma, not otherwise specified (PTCL-NOS) is a heterogeneous group of malignancies with poor outcome. Here, we identify a subgroup, PTCL-NOS <sup>SMARCB1-</sup> , which is characterized by the lack of the SMARCB1 protein and occurs more frequently in young patients. Human and murine PTCL-NOS <sup>SMARCB1-</sup> show similar DNA methylation profiles, with hypermethylation of T-cell-related genes and hypomethylation of genes involved in myeloid development. Single-cell analyses of human and murine tumors revealed a rich and complex network of interactions between tumor cells and an immunosuppressive and exhausted tumor microenvironment (TME). In a drug screen, we identified histone deacetylase inhibitors (HDACi) as a class of drugs effective against PTCL-NOS <sup>Smarcb1-</sup> . In vivo treatment of mouse tumors with SAHA, a pan-HDACi, triggered remodeling of the TME, promoting replenishment of lymphoid compartments and reversal of the exhaustion phenotype. These results provide a rationale for further exploration of HDACi combination therapies targeting PTCL-NOS <sup>SMARCB1-</sup> within the TME

DIGE Proteome Analysis Reveals Suitability of Ischemic Cardiac In Vitro Model for Studying Cellular Response to Acute Ischemia and Regeneration

Proteomic analysis of myocardial tissue from patient population is suited to yield insights into cellular and molecular mechanisms taking place in cardiovascular diseases. However, it has been limited by small sized biopsies and complicated by high variances between patients. Therefore, there is a high demand for suitable model systems with the capability to simulate ischemic and cardiotoxic effects in vitro, under defined conditions. In this context, we established an in vitro ischemia/reperfusion cardiac disease model based on the contractile HL-1 cell line. To identify pathways involved in the cellular alterations induced by ischemia and thereby defining disease-specific biomarkers and potential target structures for new drug candidates we used fluorescence 2D-difference gel electrophoresis. By comparing spot density changes in ischemic and reperfusion samples we detected several protein spots that were differentially abundant. Using MALDI-TOF/TOF-MS and ESI-MS the proteins were identified and subsequently grouped by functionality. Most prominent were changes in apoptosis signalling, cell structure and energy-metabolism. Alterations were confirmed by analysis of human biopsies from patients with ischemic cardiomyopathy

Translating cardioprotection for patient benefit: Position paper from the Working Group of Cellular Biology of the Heart of the European Society of Cardiology

Coronary heart disease (CHD) is the leading cause of death and disability worldwide. Despite current therapy, the morbidity and mortality for patients with CHD remains significant. The most important manifestations of CHD arise from acute myocardial ischaemia-reperfusion injury (IRI) in terms of cardiomyocyte death and its long-term consequences. As such, new therapeutic interventions are required to protect the heart against the detrimental effects of acute IRI and improve clinical outcomes. Although a large number of cardioprotective therapies discovered in pre-clinical studies have been investigated in CHD patients, few have been translated into the clinical setting, and a significant number of these have failed to show any benefit in terms of reduced myocardial infarction and improved clinical outcomes. Because of this, there is currently no effective therapy for protecting the heart against the detrimental effects of acute IRI in patients with CHD. One major factor for this lack of success in translating cardioprotective therapies into the clinical setting can be attributed to problems with the clinical study design. Many of these clinical studies have not taken into consideration the important data provided from previously published pre-clinical and clinical studies. The overall aim of this ESC Working Group Cellular Biology of the Heart Position Paper is to provide recommendations for optimizing the design of clinical cardioprotection studies, which should hopefully result in new and effective therapeutic interventions for the future benefit of CHD patients